Application of laser microdissection to identify the mycorrhizal fungi that establish arbuscules inside root cells.

Obligate symbiotic fungi that form arbuscular mycorrhizae (AMF; belonging to the Glomeromycota phylum) are some of the most important soil microorganisms. AMFs facilitate mineral nutrient uptake from the soil, in exchange for plant-assimilated carbon, and promote water-stress tolerance and resistance to certain diseases. AMFs colonize the root by producing inter- and intra-cellular hyphae. When the fungus penetrates the inner cortical cells, it produces a complex ramified structure called arbuscule, which is considered the preferential site for nutrient exchange. Direct DNA extraction from the whole root and sequencing of ribosomal gene regions are commonly carried out to investigate intraradical AMF communities. Nevertheless, this protocol cannot discriminate between the AMFs that actively produce arbuscules and those that do not. To solve this issue, the authors have characterized the AMF community of arbusculated cells (AC) through a laser microdissection (LMD) approach, combined with sequencing-based taxa identification. The results were then compared with the AMF community that was found from whole root DNA extraction. The AMF communities originating from the LMD samples and the whole root samples differed remarkably. Five taxa were involved in the production of arbuscules, while two taxa were retrieved inside the root but not in the AC. Unexpectedly, one taxon was found in the AC, but its detection was not possible when extracting from the whole root. Thus, the LMD technique can be considered a powerful tool to obtain more precise knowledge on the symbiotically active intraradical AMF community

Long-term impact of chemical and alternative fungicides applied to grapevine CV nebbiolo on berry transcriptome

Viticulture is one of the horticultural systems in which antifungal treatments can be extremely frequent, with substantial economic and environmental costs. New products, such as biofungicides, resistance inducers and biostimulants, may represent alternative crop protection strategies respectful of the environmental sustainability and food safety. Here, the main purpose was to evaluate the systemic molecular modifications induced by biocontrol products as laminarin, resistance inducers (i.e., fosetyl-Al and potassium phosphonate), electrolyzed water and a standard chemical fungicide (i.e., metiram), on the transcriptomic profile of ‘Nebbiolo’ grape berries at harvest. In addition to a validation of the sequencing data through real-time polymerase chain reaction (PCR), for the first-time the expression of some candidate genes in different cell-types of berry skin (i.e., epidermal and hypodermal layers) was evaluated using the laser microdissection approach. Results showed that several considered antifungal treatments do not strongly affect the berry transcriptome profile at the end of season. Although some treatments do not activate long lasting molecular defense priming features in berry, some compounds appear to be more active in long-term responses. In addition, genes differentially expressed in the two-cell type populations forming the berry skin were found, suggesting a different function for the two-cell type populations

Is Tuber brumale a threat to T. melanosporum and T. aestivum plantations?

True truffles in the genus Tuber are the most valuable ectomycorrhizal fungiand their cultivation has become widespread around the world. Competition with other ectomycorrhizal fungi and especially with undesired Tuber species, like T. brumale, can threaten the success of a truffle plantation. In this work, the competitiveness of T. brumale towards T. melanosporum and T. aestivum was assessed in a 14 year-old plantation carried out planting seedlings inoculated with these three truffle species in adjacent plots. Analyses of both truffle ectomycorrhizas and extra-radical mycelium were carried out in the transects separating the T. brumale plot from T. melanosporum and T. aestivum plots. The results confirm the competitiveness of T. brumale against T. aestivum and T. melanosporum due to its major ability to colonize the soil around its ectomycorrhizas. However, its competitiveness is limited to the transect areas and it was never found inside T. melanosporum plot. These results remark that, in presence of optimal conditions for T. melanosporum and T. aestivum, the greatest risk of contamination with T. brumale is due to wrong greenhouse activity

Intracerebral electrical stimulations of the temporal lobe: a stereo-electroencephalography study

The functional anatomy of the anteromesial portion of the temporal lobe and its involvement in epilepsy can be explored by means of intracerebral electrical stimulations. Here, we aimed to expand the knowledge of its physiological and pathophysiological symptoms by conducting the first large-sample systematic analysis of 1529 electrical stimulations of this anatomical region. We retrospectively analysed all clinical manifestations induced by intracerebral electrical stimulations in 173 patients with drug-resistant focal epilepsy with at least one electrode implanted in this area. We found that high-frequency stimulations were more likely to evoke electroclinical manifestations (p < .0001) and also provoked ‘false positive’ seizures. Multimodal symptoms were associated with EEG electrical modification (after discharge) (p < .0001). Visual symptoms were not associated with after discharge (p = .0002) and were mainly evoked by stimulation of the hippocampus (p = .009) and of the parahippocampal gyrus (p = .0212). ‘False positive seizures’ can be evoked by stimulation of the hippocampus, parahippocampal gyrus and amygdala, likely due to their intrinsic low epileptogenic threshold. Visual symptoms evoked in the hippocampus and parahippocampal gyrus, without EEG changes, are physiological symptoms and suggest involvement of these areas in the visual ventral stream. Our findings provide meaningful guidance in the interpretation of intracranial EEG studies of the temporal lobe

Extracellular polysaccharides are involved in the attachment of Azospirillum brasilense and Rhizobium leguminosarum to arbuscular mycorrhizal structures

Arbuscular mycorrhizal (AM) fungi, one of the most important component of the soil microbial community, establish physical interactions with naturally occurring and genetically modified bacterial biofertilizers and biopesticides, commonly referred to as plant growth-promoting rhizobacteria (PGPR). We have used a genetic approach to investigate the bacterial components possibly involved in the attachment of two PGPR (Azospirillum and Rhizobium) to AM roots and AM fungal structures. Mutants affected in extracellular polysaccharides (EPS) have been tested in in vitro adhesion assays and shown to be strongly impaired in the attachment to both types of surfaces as well as to quartz fibers. Anchoring of rhizobacteria to AM fungal structures may have special ecological and biotechnological significance because it may facilitate colonisation of new rhizospheres by the bacteria, and may be an essential trait for the development of mixed inocula